Figure 4. CircRHOT1 inhibits ferroptosis by sponging miR-106a-5p in breast cancer cells. (A) The potential interaction between circRHOT1 and miR-106a-5p was identified by the bioinformatic analysis using ENCORI (http://starbase.sysu.edu.cn/index.php). (B, C) The MDA-MB-231 and T47D cells were treated with the miR-106a-5p mimic or control mimic. (B) The expression levels of miR-106a-5p were measured by qPCR in the cells. (C) The luciferase activities of wild type circRHOT1 (circRHOT1 WT) and circRHOT1with the miR-106a-5p-binding site mutant (circRHOT1 MUT) were determined by luciferase reporter gene assays in the cells. (D) The MDA-MB-231 and T47D cells were treated with control shRNA or circRHOT1 shRNA. The expression of miR-106a-5p was analyzed by qPCR in the cells. (E, F) The MDA-MB-231 and T47D cells were treated with 5 mmol/L erastin, co-treated with 5 mmol/L erastin and circRHOT1 shRNA, or o-treated with 5 mmol/L erastin, circRHOT1 shRNA, and miR-106a-5p inhibitor. The cell growth was analyzed by MTT assays. (G–J) The MDA-MB-231 and T47D cells were treated control shRNA, circRHOT1 shRNA, or co-treated with circRHOT1 shRNA and miR-106a-5p inhibitor. (G, H) The levels of iron were analyzed by Iron Assay Kit. (I) The levels of ROS were measure by flow cytometry analysis in the cells. (J) The expression of GPX4, SLC7A11, and β-actin was measured by Western blot analysis in the cells. Data are presented as mean ± SD. Statistic significant differences were indicated: ** P < 0.01.